Aminoglycoside antibiotics remain the most commonly used antibiotics and the primary cause of preventable hearing loss worldwide. The impact of aminoglycoside ototoxicity has recently been aggravated by the global resurgence of tuberculosis and the increased occurrence of resistant bacteria necessitating multidrug regimens including aminoglycoside. Given the ten to 20% incidence of cochlear and vestibular disturbances associated with aminoglycoside treatment, this constitutes a major global health problem. The last decade has brought major advances in understanding the mechanisms of aminoglycoside action and designing interventions to prevent the ototoxic side effects. The proposed research is founded on past and recent discoveries from this laboratory that have led to the hypothesis of metal chelation and free radical formation by amino glycosides, and spurred by a first successful human trial demonstrating protection from gentamicin-induced hearing loss. The anticipated studies will follow new leads from preliminary experiments to delineate further the molecular mechanism of ototoxic action including the pathways of cell death and protection. Since we have recently established the adult mouse as a model for aminoglycoside ototoxicity we can combine biochemical and physiological investigations with the tools of molecular biology and the availability of mutant animals. In particular, our studies will 1. Characterize the potential contribution of lysosomal pathways to drug-induced hair cell death; 2. test the contribution of NF-kappaB mediated gene activation to cell survival; 3. analyze the regulation of the NF-kappaB pathway by signaling through phosphoinositide 3-kinase and Akt; 4. identify improved pharmacological protection based on an improved understanding of the mechanism of drug action. The results will define biochemical and molecular events involved in cell death and survival in aminoglycoside ototoxicity. The data may also help to understand other pathologies that are associated with oxidant stress, such as cisplatin ototoxicity, noise trauma and perhaps presbycusis. Optimized interventions to prevent aminoglycoside ototoxicity may serve as a basis for the translation of laboratory findings to the clinic. The attenuation or prevention of adverse effects of aminoglycosides will have far reaching implications for the continued but safe use of a family of drugs whose primary efficacy is unquestioned.